Temozolomide-Promoted MGMT Transcription Contributes to Chemoresistance by Activating the ERK Signalling Pathway in Malignant Melanoma

Abstract

Tumour cells possess a multitude of chemoresistance mechanisms, which could plausibly contribute to the ineffectiveness of chemotherapy. O⁶-methylguanine-DNA methyltransferase (MGMT) is an important effector protein associated with Temozolomide (TMZ) resistance in various tumours. To some extent, the expression level of MGMT determines the sensitivity of cells to TMZ, but the mechanism of its expression regulation has not been fully elucidated. Cultured malignant melanoma cell lines A375 and Sk-MEL28 were employed. A luciferase assay was used to detect the transcriptional activity of the MGMT promoter. Western blotting was used to compare the expression levels of phosphorylated ERK1/2 (P-ERK1/2) after TMZ treatment. Immunofluorescent staining was used to detect TMZ-induced DNA damage protein levels. The sensitivity of melanoma cells to TMZ was detected by MTT assay and animal experiments. The expression of MGMT mRNA was tested by Quantitative real-time PCR (RT-qPCR). Flow cytometry was used to measure the apoptosis of TMZ-treated cells. TMZ enhanced the transcription of MGMT through activating the ERK pathway. ERK inhibitors U0126 and vemurafenib (vMF) inhibited the TMZ induced transcription of MGMT. The expression of MGMT and p-ERK1/2 was closely related in human MM tissues. vMF increased the sensitivity of melanoma (MM) to TMZ in vitro and in vivo through downregulating MGMT and promoting the TMZ induced DNA damage in MM. TMZ-promoted MGMT transcription contributed to instinctive chemoresistance by activating the ERK signalling pathway in malignant melanoma. Our study indicates that the use of the ERK inhibitor in combination with TMZ could potentially enhance the effectiveness of clinical treatment for malignant melanoma.